The long term objective of this project is to understand the molecular genetics of obesity in humans. Obesity is the most prevalent nutrition-related problem in industrialized societies. In the U.S., approximately 1/3 of adults and 1/5 of children are obese. Prevalence rates are increasing at all ages. There is clear evidence, based on twin, adoption and segregation studies, of a substantial genetic basis for susceptibility to obesity. The specific genes, however, remain unknown. This project examines the role of human homologs of genes which produce obesity in rodents, as well as other candidate genes, by use of linkage mapping, mutation analysis and direct measurement of the protein products of these genes. Over 15,000 affected individuals (children and adults) and family members will be recruited to these studies from 13 populations (including Blacks, Caucasians, Hispanics, and American Indians) in the United States and Canada. Four of the rodent obesity genes (A [agouti], ob, fat and tub) have been cloned and shown to have human homologs. The gene products for ob (leptin), fat (carboxypeptidase E = CPE), and agouti (agouti signaling protein = ASP) have been identified and are assayable in suitable tissues/fluids. Based upon synteny relationships between human and rodent genomes, and actual gene sequences for agouti, ob, and fat, a panel of reagents has been developed for both linkage analysis and study of allelic variation in human homologs of these rodent genes. The specific aims of this project are to: 1). To collect and analyze for linkage to rodent and other candidate genes, nuclear and multiplex families in which obesity is segregating. Special emphasis is placed on acquiring families with severly affected child probands (weight > 200% average for height, age and sex) who are most likely to have strong genetic contribution to their early onset obesity. Sib pair and parametric linkage analysis will be used. 2). The results of linkage studies and of assays of gene products (protein/mRNA) - plasma leptin for OB and DB; agouti signaling protein (ASP) mRNA for agouti; carboxypeptidase E enzyme activity in saliva for fat - will be used to select individuals in whom specific genes will be examined for mutations/functionally significant allelic variation. 3). Sequence variation will be detected by SSCP and direct sequencing of coding regions and regulatory elements for these genes. The functional significance of such variation will be assessed by assaying expression levels and relevant biological activities of in vitro expressed proteins containing the predicted amino acid sequence variation. 4). Genetic screening assays will be developed for significant sequence variants and used to identify pre-obese individuals for study of the pathophysiologic mechanisms by which such variants lead to obesity.